Received: 31 October 2022 | Revised: 2 March 2023 | Accepted: 12 April 2023 DOI: 10.1002/ccr3.7282 CASE REPORT HHV8 and EBV-­negative primary effusion-­based lymphoma: A case report of a new provisional entity and review of literature Marta Canelo-­Vilaseca1 | Gustavo Tapia2 | Elisa Orna1 | Marc Sorigué1 | Isabel Granada1 | Azahara Palomar-­Muñoz3 | Ana Castillo2 | Javier Grau1 | Alba Mesa1 | Josep-­Maria Ribera1 | Juan-­Manuel Sancho1 | José-­Tomás Navarro1 1 Hematology Department, ICO-­ Hospital Germans Trias i Pujol, Josep Key Clinical Message: Carreras Leukemia Research Institute, HHV8-­ and EBV-­negative primary effusion lymphoma is an extremely rare neo- Universitat Autònoma de Barcelona, plasm involving body cavities without detectable tumor mass. It usually presents Badalona, Spain 2 Pathology Department, Hospital in elderly patients without known immunodeficiency. Compared to primary ef- Germans Trias i Pujol, Badalona, Spain fusion lymphoma, it has a better prognosis. 3 Nuclear Medicine-­PET (IDI) Primary effusion lymphoma (PEL) is a rare non-­Hodgkin lymphoma confined Department, University Hospital of exclusively to body cavities without detectable tumor masses. The term PEL-­like Bellvitge-­IDIBELL, Barcelona, Spain is an entity similar to PEL in clinical presentation but without relation to human Correspondence herpesvirus 8 (HHV8). We report a case of HHV8-­ and EBV-­negative primary José-­Tomás Navarro, Hematology effusion-­based lymphoma. Department, ICO-­Hospital Germans Trias i Pujol, Josep Carreras Leukemia Research Institute, Universitat KEYWORDS Autònoma de Barcelona, Carretera de HHV8 and EBV-­negative primary effusion-­based lymphoma, PEL, PEL-­like Canyert s/n, CP 08916 Badalona, Spain. Email: jnavarro@iconcologia.net 1 | I N T RO DU CT ION infection. In 2001, the World Health Organization (WHO) introduced the category of PEL only for HHV8-­related pri- Primary effusion lymphoma (PEL) is a rare aggressive mary lymphomatous effusions. In 1996, Hermine et al.4 B-­cell neoplasm that presents with serous malignant ef- reported the first HHV8-­negative lymphomatous effusion fusions without detectable tumor-­associated mass.1 It is in an HIV-­seronegative patient. Since then, more than 60 related to HHV8 infection and mainly affects immunocom- HHV8-­unrelated effusion-­based lymphomas (EBL) have promised patients such as HIV-­infected individuals, recip- been reported in the literature, most of them in Japan, ients of solid-­organ transplantation, or elderly patients.3 and terms as EBL and PEL-­like lymphomas have been Moreover, coinfection with Epstein–­Barr virus (EBV) is used. Recently, the International Consensus Classification frequent. The term “primary effusion lymphoma” was first (ICC) of Mature Lymphoid Neoplasms has recognized described in the literature in 1996 by Nador et al.3 to de- these cases as a new provisional entity named HHV8 fine malignant lymphomatous effusions related to HHV8 and EBV-­negative primary effusion-­based lymphoma.2 This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made. © 2023 The Authors. Clinical Case Reports published by John Wiley & Sons Ltd. Clin Case Rep. 2023;11:e07282.  wileyonlinelibrary.com/journal/ccr3   | 1 of 6 https://doi.org/10.1002/ccr3.7282 2 of 6 |    CANELO-­VILASECA et al. Moreover, WHO Classification, 5th edition, has also re- cell/μL with 55% of atypical cells, protein 49.6 g/L, LDH cently recognized this entity as fluid overload-­associated 4034 U/L, ADA 84.2 U/L). Malignant-­appearing cells were large B-­cell lymphoma.5 detected in the cytospin preparation. Microbiological We report a 72-­year-­old patient that presented with tests were negative including mycobacteria culture and pericardial effusion with no evidence of tumor mass. multiplex polymerase chain reaction (PCR) for herpesvi- Morphologic, immunophenotypic, and molecular stud- rus (VHS1, VHS2, VVZ, CMV, and VH6). A whole-­body ies of the tumor cells of the pericardial effusion were positron emission tomography (PET) scan showed mod- performed. Cytological analysis showed large cells with erately hypermetabolic pericardial and pleural layers and irregular nuclei, prominent nucleoli, and vacuolized cy- right tonsillar enlargement (Figure 1B). Tonsil excisional toplasm. Immunophenotype showed expression of CD45, biopsy showed reactive follicular hyperplasia and a bone CD19, and CD20 without detection of surface immuno- marrow biopsy showed no evidence of involvement by globulin. The tumor cells were negative for LANA-­1 and lymphoma. Serologic tests were negative for HIV, hepa- EBER. Finally, PEL-­like was diagnosed. To date, clinical titis C virus (HCV), and hepatitis B virus (HBV); whereas and pathologic features of HHV8-­unrelated, effusion-­ immunoglobulin G for EBV was positive, indicating a based lymphomas (EBL) are poorly understood. We prior infection. report herein a case of the new entity HHV8-­ and EBV-­ Cytological evaluation demonstrated large atypical negative primary effusion-­based lymphoma, presenting in cells variable in size and nuclei, ranging from round to a Caucasian woman, and review the literature. more irregular shape with prominent nucleoli and abun- dant basophilic cytoplasm with vacuoles (Figure 2A). Moreover, some mitotic figures were seen (Figure 2B). 2 | C A S E RE PORT In flow cytometric analysis, these cells were positive for CD45, CD19, CD10, CD20, and CD38 and lacked surface We report the case of a 72-­year-­old Caucasian woman, with immunoglobulin expression (Figure 3). A formalin-­fixed a medical history of hypertension, hypothyroidism, and paraffine-­embedded cell block was performed for mor- mild obesity, who was admitted to the hospital because of phology, immunocytochemistry (IHC), and fluorescence progressive dyspnea and appearance of bilateral leg oede- in situ hybridization (FISH) evaluation. IHC study showed mas above the ankles. No weight loss or concomitant fever positivity for CD20 (Figure 4A) and PAX5 and negativity was reported. Physical examination revealed distant heart for CD3, CD10, CD30, MUM-­1, BCL-­6, BCL-­2, c-­myc, TdT, sounds. The liver, spleen, and peripheral lymph nodes and myeloperoxidase. Neither BCL-­6, BCL-­2, nor C-­MYC were not palpable. Laboratory data on admission was re- gene translocation was detected by FISH. Moreover, EBV-­ markable for a B-­type natriuretic peptide level of 1117 pg/ encoded RNA (EBER) was negative (Figure 4B), and pro- mL (normal value: <250 pg/mL). Complete blood count, tein latency-­associated nuclear antigen 1 (LANA-­1) was renal and hepatic function were within normal values and negative on IHC (Figure 4D). Positive EBER and LANA-­1 the serum lactate dehydrogenase (LDH) was not elevated. pictures have been included as control images (Figure 4C A chest X-­ray showed cardiomegaly (Figure 1A) and tran- and Figure 4E, respectively). High Ki67 expression was sthoracic echocardiogram revealed pericardial effusion. observed (>80%). Cytogenetic findings showed a complex The patient was admitted to the cardiac care unit, where karyotype (Figure 5). pericardiocentesis was successfully carried out, yielding All these findings pointed toward a diagnosis of PEL-­ 800 mL of serohematic fluid. The pericardial liquid analy- like or HHV8 and EBV-­negative primary EBL according sis showed features of exudative effusion (glucose <5 mg/ to the ICC.2 The patient was treated with R-­CHOP (rit- dL, red blood cells 15,871 cell/μL, white blood cells 850 uximab, cyclophosphamide, doxorubicin, vincristine, and (A) (B) F I G U R E 1 (A) Chest x-­ray showing cardiomegaly. (B) Moderate uptake in the pleura and pericardium in 2-­deoxy-­2-­ (18F)fluoro-­D-­glucose positron emission tomography/computed tomography with (FDG-­PET/CT). CANELO-­VILASECA et al.    | 3 of 6 (A) (B) F I G U R E 2 Morphologic appearance of malignant cells in pericardial fluid. (A) Large atypical lymphoid cells with irregular nuclei, prominent nucleoli, and basophilic cytoplasm in the pericardial effusion (cytospin preparation, May-­Grünwald-­Giemsa, ×500). (B) Showing a cell in mitosis (cytospin preparation, May-­Grünwald-­Giemsa, ×1000). F I G U R E 3 Flow cytometric analysis shows abnormal B cell population, with high intensity for both CD45 and CD19, lacked surface immunoglobulin expression, and positive for CD10 and CD38. prednisone) for six cycles achieving complete remission HHV8-­positive PEL is a recognized entity in the current evaluated by PET scan. At the time of this report, the pa- WHO classification1; but HHV8 and EBV-­negative PEBL tient remains in complete response for 16 months. has been recently suggested as a provisional entity by the ICC.2 Our case is of particular interest because of the 3 | DI S C USSION Caucasian origin of the patient as most of the published data are case series reported from Japan.6 Despite its dif- PEL-­like or HHV8 and EBV-­negative PEBL is a rare B-­cell ferent origin, the other characteristics are similar to those neoplasm that typically presents as a malignant serous ef- described in the Japanese cases. Its prevalence is unknown fusion in non-­immunocompromised patients. In the last and its diagnosis is challenging (especially because it has decade, several reports of HHV8-­negative effusion lym- not been described in the current WHO classification). phomas have been published demonstrating remarkably HHV8 and EBV-­negative PEL are considered to similar cytomorphologic characteristics to PEL but dif- have a distinct etiopathogenic mechanism than HHV8-­ ferences in immunophenotype, demographics, response positive PEL. Different theories have emerged with no to treatment, and clinical outcome; suggesting that PEL-­ conclusive data to date. Kobashi et al.7 suggested in like is a distinct entity. As mentioned before, to date only 2007 that immunosenescence related to age could play 4 of 6 |    CANELO-­VILASECA et al. (A) (B) (C) (D) (E) F I G U R E 4 Cell-­block specimen of PEL-­like from patients' pericardial effusion stained with anti-­CD20 (A), EBV-­encoded small RNA (EBERs) (B), positive EBER control (C), LANA-­1 (D), and positive LANA-­1 control (E). a major role, as most of the patients were older (median Although this last hypothesis needs further confirma- age of 70 years) compared with PEL individuals. In the tion and not all the authors agree on that.6 same line, Ohshima et al.8 concluded that multi-­step ge- Tumor cells in HHV8 and EBV-­negative PEL express nomic abnormalities found in PEL-­like cases could be pan-­B-­cell markers and they are negative for plasma cell related to age. In their study, most of the patients were markers in contrast with PEL. CD45 antigen expression elderly men with all cases showing complex karyotypes is present in both neoplasms indicating a hematological but without identification of a driver genomic abnor- derivation of the effusion cells.10 mality. Suggested mechanisms of lymphomagenesis in According to the reported series, the site of the effu- HHV8 and EBV-­negative PEL include impaired immune sion varies but the most common location was pleural, surveillance and dysregulatory hemopoietic pathway. followed by ascites, and being the pericardial one the less Unlike PEL, in which HHV8 acts as an exogenous stim- common.7 Nonetheless, approximately 40% of the cases ulus for local B-­cell expansion, no viral mechanism has showed multiple body cavity lesions including pericardial yet been identified in PEL-­like lymphomas. In our case, in all of them.6 In our case, probably both pericardial and Epstein–­Barr virus-­encoded small RNA in situ hybrid- pleural effusions had a neoplastic involvement according ization was negative but the patient had seropositivity to images from PET scan (Figure 2B). Nonetheless, the for EBV. In the literature, around one-­third of cases of reason why this B-­cell neoplasm is exclusively confined PEL-­like lymphomas are infected with EBV.9 However, to serous membranes without developing tumor masses is whether EBV is simply a prior infection or a trigger of still non-­explained. PEL-­like lymphoma remains unclear and further inves- Diffuse Large B-­cell lymphoma associated with chronic tigation is needed. Regarding HCV infection in PEL-­like inflammation is another B-­cell neoplasm that also arises lymphomas, there is a slightly higher prevalence of its in the serous membranes but unlike our case, the patho- infection among these patients (30%–­40% of patients). genic mechanism and localization of this entity are well Association between HCV and B-­cell non-­Hodgkin lym- understood. Chronic inflammation causes local immu- phoma is well-­reported in the literature. Thus, it has nosuppression permitting EBV-­infected memory B-­cells been suggested that fluid overload related to cirrhosis to evade immunosurveillance, accumulate genetic alter- due to HCV infection plays a role in PEL-­like lympho- ations, and evolve to lymphoma.11 mas. Moreover, an underlying medical condition leading There is no standard treatment for this entity due to fluid overload has been observed too in almost one-­ to the small number of cases although the majority of half of the patients. 7 The ICC also recognizes chronic se- them are treated with chemotherapy including anth- rosal stimulation by fluid overload in its pathogenesis.2 racycline and rituximab (as tumor cells express CD20). CANELO-­VILASECA et al.    | 5 of 6 FIGURE 5 Karyotyping results of the abnormal cells demonstrate a complex karyotype. In a retrospective study conducted by Kaji et al.6 86% of and EBV-­negative PEL are usually present in elderly pa- patients received CHOP-­like regimens. Other systemic tients without known immunodeficiency. Distinguishing treatments consisted of rituximab in monotherapy or in PEL-­like from PEL is important because the better re- combination with etoposide. Perhaps, due to the intro- sponse to treatment and outcomes are achieved with duction of rituximab, the prognosis seems to be better in standard therapies. Further investigation is needed to PEL-­like lymphomas.12 In this study conducted in Japan,6 better understand this entity. To our knowledge, few the 2-­year overall survival was 84.7% (95% confidence in- cases of this newly recognized entity have been described terval, 71.3–­92.1) compared to the 1-­year overall survival in Caucasians. of 30% in PEL. Moreover, there are some cases that were successfully treated only with drainage of the effusion.13 AUTHOR CONTRIBUTIONS Its survival might be conditioned by the advanced age at Marta Canelo-­Vilaseca: Formal analysis; investigation; diagnosis and patients' comorbidity.14 writing –­ original draft; writing –­ review and editing. Gustavo Tapia: Data curation; investigation; resources; validation. Elisa Orna: Methodology. Marc Sorigué: 4 | CO N C LUSION Data curation; investigation. Isabel Granada: Data cura- tion; investigation. Azahara Palomar-­Muñoz: Data cu- In summary, HHV8 and EBV-­negative PEL is an ex- ration; investigation. Ana Castillo: Investigation. Javier tremely rare neoplasm involving body cavities without Grau: Data curation. Alba Mesa: Data curation. Josep-­ detectable tumor mass, characterized by atypical blastic Maria Ribera: Supervision; validation. Juan-­Manuel cells expressing pan-­B-­cell markers. Unlike PEL, HHV8 Sancho: Supervision; validation. Jose-­Tomas Navarro: 6 of 6 |    CANELO-­VILASECA et al. 5. Alaggio R, Amador C, Anagnostopoulos I, et al. The 5th edi- Investigation; project administration; supervision; valida- tion of the World Health Organization Classification of tion; writing –­ review and editing. Haematolymphoid Tumours: lymphoid neoplasms. Leukemia. 2022;36(7):1720-­1748. FUNDING INFORMATION 6. Kaji D, Ota Y, Sato Y, et al. Primary human herpesvirus No funding was requested/provided for this article. 8-­negative effusion-­based lymphoma: a large B-­cell lymphoma with favorable prognosis. Blood Adv. 2020;4(18):4442-­4450. CONFLICT OF INTEREST STATEMENT 7. Kobayashi Y, Kamitsuji Y, Kuroda J, et al. Comparison of human herpes virus 8 related primary effusion lymphoma with human The authors declared no competing interest. herpes virus 8 unrelated primary effusion lymphoma-­like lym- phoma on the basis of HIV: report of 2 cases and review of 212 DATA AVAILABILITY STATEMENT cases in the literature. Acta Haematol. 2007;117(3):132-­144. The authors declare that data supporting the findings of 8. Ohshima K, Ishiguro M, Yamasaki S, et al. Chromosomal and this study are available within the article. comparative genomic analyses of HHV-­8-­negative primary ef- fusion lymphoma in five HIV-­negative Japanese patients. Leuk CONSENT Lymphoma. 2002;43(3):595-­601. 9. Alexanian SJ, Said J, Lones M, Pullarkat ST. KSHV/HHV8-­ Written informed consent was obtained from the patient negative effusion-­based lymphoma, a distinct entity associated to publish this report in accordance with the journal's pa- with fluid overload states. Am J Surg Pathol. 2013;37:241-­249. tient consent policy. 10. Shimada K, Hayakawa F, Kiyoi H. Biology and management of primary effusion lymphoma. Blood. 2018;132(18):1879-­1888. ORCID 11. Loong F, Chan ACL, Ho BCS, et al. 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